In a cooperative effort between the University of Texas and the Upjohn Company it is proposed to establish the biosynthetic precursors for trimycin, determine the precise manner in which trimycin reacts with DNA, and define the biological consequences of DNA damage in human cell lines. Trimycin, an antitumor antibiotic produced by Streptomyces zelensis, is probably the most potent antitumor agent discovered to date and appears to have a unique structure and mechanism of action. Biosynthetically we propose to determine the building blocks for trimycin and elucidate the manner in which the cyclopropane ring is formed by using radioactive (3H and 14C) and stable isotope (13C) techniques. We propose the twisted structure of trimycin binds within one of the grooves of DNA and alkylates via a spirocyclopropane substituted indole-quinone. The structure of the trimycin-DNA adduct will be determined by either isolation of a drug-nucleotide adduct or by 1H and 13C-NMR studies on 13C-enriched trimycin attached to DNA. Using established techniques with repair proficient and deficient human cell lines we will determine the biological consequences of reaction of trimycin with DNA and the repair pathways involved.